The long term objective is to discover which aspects of differential FSH and LH release are regulated directly at the level of the anterior pituitary gland itself. Basically, two different in vitro setups will be used: (a) a dynamic perifusion apparatus using dispersed cells; (b) a dynamic perifusion apparatus using quartered pituitaries. Under selected experimental conditions a more conventional static, dispersed pituitary cell culture will also be employed. These experimental setups will permit contrast of dynamic vs static input-output relations as well as contrast of dynamic control in the absence (dispersed) and presence (quartered pituitaries) of undisrupted cell-cell connections. Measurements to be made include LH and FSH in the medium and in the pituitary gland, as well as pituitary GnRH receptor levels. The first aim of these studies is to examine FSH and LH secretory rates in response to variations in intensity, duration and/or frequency of GnRH input, frequently cited as accounting for differences in the secretion rates of the two gonadotropins. Initial donors will be intact males and intact females at each cycle stage. We will then determine the effect of in vitro treatment with gonadal steroids and peptide on FSH and LH secretion in basal and GnRH-stimulated pituitaries. The literature is contradictory on the site(s) of gonadal negative feedback; our experimental model will enable us to probe for interactions between these feedback factors and a dynamic GnRH signal, at the level of the pituitary gland itself. Next we will examine the effect of in vivo increases or decreases in gonadal feedback in donor males and females on the in vitro behavior of their pituitaries, when cell-cell connections are either disrupted or retained. Finally, we will probe whether the anterior pituitary gland itself is an important site of interaction between the CRF-ACTH-corticoid axis and gonadotropin secretion rates. In vivo, castrated male rats pretreated with cortisol show a suppressed LH response to GnRH, but the FSH secretory response to GnRH is normal, and pituitary FSH stores are increased. These findings were important, because they suggested that glucocorticoids could be an important probe in the search for the site of differential FSH regulation. We were unable to replicate this finding in a standard static, dispersed pituitary cell culture. Can we recover the LH suppression and FSH enhancement treating quartered pituitaries in perifused culture with glucocorticoids?